Stem/progenitor cell transplantation-based replacement of astrocytes is a novel and potentially powerful therapeutic strategy for treating CNS diseases such as traumatic spinal cord injury (SCI). Towards this goal, we propose to test transplantation of human induced Pluripotent Stem (iPS) cell-derived astrocytes (hiPSAs) for mitigating SCI-induced neuropathic pain. A major portion of SCI patients suffer from neuropathic pain, resulting in often long-term physical and psychological burdens. Effective treatments for this debilitating outcome of SCI are currently lacking; therefore, novel and robust therapeutic approaches are urgently needed. Following SCI, chronic dysregulation of extracellular glutamate homeostasis plays a key role in persistent central hyperexcitability of superficial dorsal horn (DH) neurons that mediate pain neurotransmission, leading to neuropathic pain. Astrocytes are the principal cell type that expresses the glutamate transporter, GLT1, which is responsible for the vast majority of glutamate uptake in the intact CNS, particularly in spinal cord. In our rodent model of cervical contusion SCI that results in persistent neuropathic pain, we find significant and long-lasting reductions in GLT1 expression and functional GLT1-mediated glutamate uptake in cervical DH. iPS cells are a novel and clinically-relevant source for homogeneous derivation of mature cell types in large quantities for applications such as transplantation, potentially in an autologous fashion from the eventual patient recipient. The use of iPS cell transplantation to deliver astrocytes represents an exciting strategy that has not been extensively studied to date, particularly with respect both to targeting key astrocyte functions such as glutamate uptake at a mechanistic level and to addressing neuropathic pain. Importantly, a majority of human spinal cord trauma cases affect cervical regions and are of the contusion/compression type, urgently calling for the assessment of disease etiology and treatment of neuropathic pain specifically in models of cervical contusion SCI to provide relevance to the patient population. In Aim 1a, we will determine whether hiPSA transplantation can reverse several forms of established neuropathic pain-related behavior in a rat model of cervical contusion. Importantly, we will assess motivational and affective components of pain using an operant paradigm. In Aim 1b, we will determine whether hiPSAs can reduce hyperexcitability of DH pain neurons using whole-cell patch clamp recording in an intact ex vivo preparation. In Aim 2a, we will examine the ability of hiPSA transplants to express GLT1 and to restore GLT1- mediated glutamate uptake levels in DH after cervical contusion. We will also engineer hiPSAs to overexpress GLT1 to enhance their therapeutic potential. In Aim 2b, we will examine whether hiPSAs can alter the DH microglia/macrophage response that plays a central role in hyperexcitability and chronic pain. We will assess whether hiPSAs can shift this response after SCI away from a M1 pro-inflammatory phenotype (and toward a M2 reparative phenotype) as an additional - and potentially powerful - mechanism of hiPSA transplant action.